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The Structure of Ca2+ Sensor Case16 Reveals the Mechanism of Reaction to Low Ca2+ Concentrations

Leder, Lukas and Stark, Wilhelm and Freuler, Felix and Marsh, May and Meyerhofer, Marco and Stettler, Thomas and Mayr, Lorenz and Britanova, Olga V and Strukova, Lydia A and Chudakov, Dmitriy M and Souslova, Ekaterina A (2010) The Structure of Ca2+ Sensor Case16 Reveals the Mechanism of Reaction to Low Ca2+ Concentrations. Sensors, 10 (9). pp. 8143-8160. ISSN 1424-8220

Abstract

Here we report the first crystal structure of a high-contrast genetically encoded circularly permuted green fluorescent protein (cpGFP) - based Ca2+ sensor Case16 in presence of low Ca2+ concentration. The obtained structure revealed that the actual positioning of the chromophore environment within Case16 was fixed at the first stage of the Ca2+-dependent response when only two out of four Ca2+-binding pockets of calmodulin (CaM) are occupied with Ca2+ ions. It was demonstrated that in such a “half Ca2+-bound state” Case16 was characterized by incomplete interaction of its CaM-/M13-domains. Another structure reported herein is the crystal structure of the related Ca2+ sensor Case12 at saturating Ca2+ concentration. Based on this structure we postulate that cpGFP-based Ca2+ sensors can form non-functional homodimers where the CaM-domain of one sensor molecule symmetrically binds to the M13-peptide of the partner sensor molecule. Studying the dependence of Case12 and Case16 behavior upon addition of high concentrations of free CaM or M13-peptide revealed that the latter effectively blocks the fluorescent response of the sensor. We speculate that the demonstrated intermolecular interaction with endogenous substrates and homodimerization are the key reasons impeding the performance of this type of Ca2+ indicators in neurons and other living systems.

Item Type: Article
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Date Deposited: 13 Oct 2015 13:16
Last Modified: 13 Oct 2015 13:16
URI: https://oak.novartis.com/id/eprint/3315

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